Campylobacter jejuni is the leading cause of human gastroenteritis worldwide with over 500 million cases annually. Chemotaxis and motility have been identified as important virulence factors associated with C. jejuni colonisation. Methyl-accepting chemotaxis proteins (MCPs), also termed transducer-like proteins (Tlps), serve as sensors in C. jejuni chemotactic signalling. Tlp receptors are responsible for sensing the external environment for bacterial movement to or away from a chemical gradient or a stimulus. The focus of this study is to characterise the C. jejuni Tlp3 chemoreceptor (cj1564) by determining its ligand-binding specificity, signalling pathway and function, through assessing the effect of tlp3 mutation in vitro using cell culture and in vivo using avian hosts. The purified periplasmic domain of Tlp3 was used to identify ligands associated with the protein (namely, isoleucine, asparagine, aspartic acid, glutamic acid and arginine as well as succinic acid, glucosamine, tricine, malic acid, fumaric acid, fructose, thiamine, purine, [alpha]-ketoglutarate and galactose). An ELISA based plate assay was used as a pre-screening tool to identify multiple potential ligands and the obtained results were further verified by using array technology.